GSBS Student Publications


Nitric oxide-mediated inhibition of Hdm2-p53 binding

UMMS Affiliation

Graduate School of Biomedical Sciences; Department of Biochemistry and Molecular Pharmacology; Department of Cell Biology



Document Type


Medical Subject Headings

Amino Acid Sequence; Binding Sites; Cysteine; Dithiothreitol; Enzyme-Linked Immunosorbent Assay; Glutathione; Glutathione Transferase; Humans; Molecular Sequence Data; Mutagenesis, Site-Directed; Mutation; Neoplasm Proteins; *Nuclear Proteins; Protein Binding; Protein Conformation; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-mdm2; Recombinant Fusion Proteins; Sequence Homology, Amino Acid; Triazenes; Tumor Suppressor Protein p53


Life Sciences | Medicine and Health Sciences


It has become increasingly evident that nitric oxide exerts its effects, in part, by S-nitrosylation of cysteine residues. We tested in vitro whether nitric oxide may indirectly control p53 by S-nitrosylation and inactivation of the p53 negative regulator, Hdm2. Treatment of Hdm2 with a nitric oxide donor inhibits Hdm2-p53 binding, a critical step in Hdm2 regulation of p53. The presence of excess amounts of cysteine or dithiothreitol blocks this inhibition of binding. Moreover, nitric oxide inhibition of Hdm2-p53 binding was found to be reversible. Sulfhydryl sensitivity and reversibility are consistent with nitrosylation. Finally, we have identified a critical cysteine residue that nitric oxide modifies to disrupt Hdm2-p53 binding. This cysteine is proximal to the Hdm2-p53 binding interface and is conserved across species from zebrafish to humans. Mutation of this residue from a cysteine to an alanine does not interfere with binding but rather eliminates the sensitivity of Hdm2 to nitric oxide inactivation.

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Citation: Biochemistry. 2002 Nov 19;41(46):13570-4.

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